Experimental and clinical observations have supported the concept that the hypothalamus plays a key role in the regulation of adenohypophysial corticotropic cells secretory functions. Although over 40 years ago, Guillemin, Rosenberg and Saffran and Schally independently demonstrated the presence of factors in hypothalamus which would increase the rate of ACTH secretion by the pituitary gland incubated in vitro or maintained in an organ culture, a physiologic corticotropin releasing factor (CRF) was not characterized until ovine CRF (oCRF) was characterized in 1981. It was disclosed in U.S. Pat. No. 4,415,558, as having the amino acid sequence (SEQ ID NO:1):
H-Ser-Gln-Glu-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu-Val-L eu-Glu-Met-Thr-Lys-Ala-Asp-Gln-Leu-Ala-Gln-Gln-Ala-His-Ser-Asn-Arg-Lys-Leu- Leu-Asp-Ile-Ala-NH.sub.2. PA0 H-Ser-Glu-Glu-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu-Val-L eu-Glu-Met-Ala-Arg-Ala-Glu-Gln-Leu-Ala-Gln-Gln-Ala-His-Ser-Asn-Arg-Lys-Leu- Met-Glu-Ile-Ile-NH.sub.2. The formula of human CRF was subsequently determined to be the same as that of rCRF. The compound is often referred to as r/hCRF and is covered in U.S. Pat. No. 4,489,163. PA0 H-Asn-Asp-Asp-Pro-Pro-Ile-Ser-Ile-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Asn-Met-I le-Glu-Met-Ala-Arg-Ile-Glu-Asn-Glu-Arg-Glu-Gln-Ala-Gly-Leu-Asn-Arg-Lys-Tyr- Leu-Asp-Glu-Val-NH.sub.2 ; it is sometimes referred to as suckerfish (sf) urotensin or sfUI. Its purification and characterization are described in an article by Lederis et al., Science Vol. 218, No. 4568, 162-164 (Oct. 8, 1982). The homolog, carp urotensin, was obtained from Cyprinus carpio and has the amino acid sequence (SEQ ID NO:5): PA0 H-Asn-Asp-Asp-Pro-Pro-Ile-Ser-Ile-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Asn-Met-I le-Glu-Met-Ala-Arg-Asn-Glu-Asn-Gln-Arg-Glu-Gln-Ala-Gly-Leu-Asn-Arg-Lys-Tyr- Leu-Asp-Glu-Val-NH.sub.2. Another urotensin homolog having the following amino acid sequence (SEQ ID NO:6): PA0 H-Ser-Glu-Glu-Pro-Pro-Met-Ser-Ile-Asp-Leu-Thr-Phe-His-Met-Leu-Arg-Asn-Met-I le-His-Arg-Ala-Lys-Met-Glu-Gly-Glu-Arg-Glu-Gln-Ala-Leu-Ile-Asn-Arg-Asn-Leu- Leu-Asp-Glu-Val-NH.sub.2 was later isolated from the urophyses of Hippoglossoides elassodon or Flathead (Maggy) Sole; it is sometimes referred to as Maggy urotensin. Synthetic UIs have been found to also stimulate ACTH and .beta.-endorphin activities in vitro and in vivo and to have many of the same general biological activities of CRFs and sauvagine.
Although originally isolated and characterized on the basis of its role in this hypothalamopituitary-adrenal (HPA) axis, CRF has been found to be distributed broadly throughout the central nervous system as well as in extraneural tissues, such as the adrenal glands, placenta and testes, where it may also act as a paracrine regulator or a neurotransmitter. Moreover, the likely involvement of CRF in affective disorders, such as anxiety, depression, alcoholism and anorexia nervosa, and in modulating reproduction and immune responses suggests that changes in CRF expression may have important physiological and pathophysiological consequences. For example, perturbations in the regulatory loops comprising the HPA axis often produce chronically elevated levels of circulating glucocorticoids; such patients display the physical hallmarks of Cushing's syndrome, including truncal obesity, muscle-wasting, and reduced fertility.
In addition to its role in mediating activation of the hypothalamic-pituitary-adrenal, CRF has also been shown to modulate autonomic and behavioral changes, some of which occur during the stress response. Many of these behavioral changes have been shown to occur independently of HPA activation in that they are not duplicated by dexamethasone treatment and are insensitive to hypophysectomy. In addition, direct infusion of CRF into the CNS mimics autonomic and behavioral responses to a variety of stressors. Because peripheral administration of CRF or a CRF antagonist fails to affect certain of these changes, it appears that CRF exhibits a direct brain action with respect to such functions, which include appetite suppression, increased arousal and learning ability. However, CRF antagonists given peripherally attenuate stress-mediated increases in ACTH secretion, and when delivered into the cerebral ventricles can mitigate stress-induced changes in autonomic activity and behavior.
As a result of the extensive anatomical distribution and multiple biological actions of CRF, this regulatory peptide is believed to be involved in the regulation of numerous biological processes. CRF has also been implicated in the regulation of inflammatory responses. Although it has been observed that CRF plays a pro-inflammatory role in certain animal models, CRF appears to suppress inflammation in others by reducing injury-induced increases in vascular permeability.
In about 1981, a 40-residue amidated peptide generally similar to CRF was isolated from the skin of the South American frog Phyllomedusa sauvagei; it is referred to as sauvagine. It was characterized by Erspamer et al. and was described in Regulatory Peptides, Vol. 2 (1981), pp. 1-13. Sauvagine has the amino acid sequence (SEQ ID NO:2): pGlu-Gly-Pro-Pro-Ile-Ser-Ile-Asp-Leu-Ser-Leu-Glu-Leu-Leu-Arg-Lys-Met-Ile-G lu-Ile-Glu-Lys-Gln-Glu-Lys-Glu-Lys-Gln-Gln-Ala-Ala-Asn-Asn-Arg-Leu-Leu-Leu- Asp-Thr-Ile-NH.sub.2. When given intravenously (iv), sauvagine and oCRF have been reported to cause vasodilation of the mesenteric arteries so as to lower blood pressure in mammals and also in stimulating the secretion of ACTH and .beta.-endorphin. However, when administered intracerebroventricularly (icv), there is an elevation of heart rate and mean arterial blood pressure, which are secondary to activation of the sympathetic nervous system.
Rat CRF (rCRF) was isolated, purified and characterized in about 1982-1983 as a hentetracontapeptide having the amino acid sequence (SEQ ID NO:3):
At about the same time, two homologous polypeptides were isolated from the urophyses of different species of fish. These isolated peptides were generally homologous to CRF, i.e. about 54% homology, and were termed Urotensin I (UI). Catostomus commersoni (white sucker or suckerfish) UI is a polypeptide having the amino acid sequence (SEQ ID NO:4):
Since the discovery of the original discoveries of CRFs in mammals and urotensins in fish, CRFs have now been shown to exist in other animal species. For example, fish CRF was found to be a 41-residue peptide having high homology to r/hCRF; it has the amino acid sequence (SEQ ID NO:7): H-Ser-Glu-Glu-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu-Val- Leu-Glu-Met-Ala-Arg-Ala-Glu-Gln-Leu-Ala-Gln-Gln-Ala-His-Ser-Asn-Arg-Lys-Met -Met-Glu-Ile-Phe-NH.sub.2. Synthetic fish CRF (fCRF) stimulates ACTH and .beta.-endorphin activities in vitro and in vivo and has similar biological activities to mammalian CRFs. Because of the high homology between fCRF and r/hCRF, it is thought that other mammalian hormones may exist which would be the counterparts of urotensin and/or sauvagine.